The ability of stem cells to develop into a large variety of different cell types holds great promise for treatment of many human diseases. Furthermore, the control of the growth of stem cells and other unspecified cells is critical to prevent the development of cancer. Underlying these processes are fundamental aspects of cell biology involved in signaling between cells, and the control of cell orientation. This proposal seeks to address these issues by studying the fundamental regulation of stem cell specification, differentiation, and polarity as controlled by the CLV signaling pathway in Arabidopsis. The long-terms goals are to understand mechanism by which the delicate balance between stem cell proliferation and differentiation is maintained, and the mechanism by which CLV signaling regulates these processes. The CLV signaling pathway is a key factor -the loss of any component in this pathway leads to massive accumulation of stem cells at the expense of differentiation. Recent findings in the field have uncovered novel components of the signaling pathway, and provided key insights into the mechanism of CLV signaling. This proposal will explore the role of the POL/PLL1 phosphatases in signaling and the establishment of cell polarity. Receptor sub-cellular localization will be addressed, along with receptor co-localization and interaction with putative downstream effectors. Preliminary evidence of receptor association with lipid rafts will be rigorously tested. The role of cofilin, which binds to the CLV1 receptor-kinase, will tested through a combination of genetic, localization and protein interaction approaches. The structure/function and possible maturation of the putative ligand CLV3 will be investigated. A BY-2 cell culture system will be developed to integrate these different experimental lines in a way that addresses receptor activation and mechanism of signaling. The results of these experiments will provide major steps forward in understanding these key developmental processes. [unreadable] [unreadable] [unreadable]